DNA repair is essential to maintaining cellular health and preventing tumorigenesis. This application seeks funding to study the normal function and regulation of the non-homologous end joining (NHEJ) factor DNA-dependent protein kinase catalytic subunit (DNA-PKcs), and its substrate Artemis, during lymphocyte development and lymphomagenesis to elucidate how mutations in these proteins lead to severe combined immunodeficiency (SCID) and lymphomas. V(D)J recombination, the repair of programmed double-strand breaks (DSBs) in the Immunoglobulin (Ig) loci, is required for normal B cell development from lymphoid progenitors. DNA repair during V(D)J recombination requires the core NHEJ factors (Ku70, Ku80, Lig4, XRCC4, and XLF), and vertebrate specific DNA-PKcs and Artemis. While DNA-PKcs and Artemis are not strictly required for end-ligation, they are required for end processing such as opening hairpin-sealed coding ends during V(D)J recombination. Therefore, DNA-PKcs or Artemis null mutations result in isolated SCID, without the neuronal defects found with core-NHEJ deficiency. DNA-PKcs belongs to the PI3 Kinase related kinase family, which also includes ATM. ATM and DNA-PKcs share many common substrates, including DNA- PKcs itself. To determine whether DNA-PKcs protein has a role in DNA repair beyond its kinase activity, we knocked in the D3922A kinase dead (KD) mutation in murine DNA-PKcs (Prkdc). In contrast to the normal development of DNA-PK null mice, DNA-PKcsKD/KD, but not DNA-PKcs+/KD, mice died during late embryonic development with severe genomic instability, similar to core-NHEJ deficient mice (eg.Lig4). Surprisingly DNA- PKcsKD/KD mice were defective in end-ligation, but not in end processing (hairpin opening). Despite the severe genomic instability, DNA-PKcsKD/KDp53-/- mice have less lymphoma than the DNA-PKcs-/-p53-/- mice. Based on these and other findings, we hypothesize that the DNA-PKcs protein structure suppresses lymphomagenesis and supports normal lymphocyte development by regulating NHEJ, alternative-end- joining and oncogenic translocations. To test this, we propose to investigate (Aim 1) the noncatalytic function of DNA-PKcs in NHEJ and hairpin opening; (Aim 2) the noncatalytic function of DNA-PKcs in CSR and A-EJ; and (Aim 3) the role of DNA-PKcs in B cell lymphomagenesis. Together, these studies will determine the physiological function of the DNA-PKcs protein during DNA double strand break repair and lymphocyte development and how mutations in DNA-PKcs and its substrates leads to primary immunodeficiency and cancer.